MM165 - A Small Hybrid Molecule Modulates the Kynurenine Pathway and Attenuates Lipopolysaccharide-Induced Memory Deficits and Inflammation.

Cognitive dysfunctions are now recognized as core symptoms of various psychiatric disorders e.g., major depressive disorder. Sustained immune activation may leads to cognitive dysfunctions. Proinflammatory cytokines shunt the metabolism of tryptophan towards kynurenine and quinolinic acid may accumulate at toxic concentrations. This acid triggers an increase in neuronal nitric oxide synthase function and promotes oxidative stress. The searching for small molecules that can regulate tryptophan metabolites produced in the kynurenic pathway has become an important goal in developing treatments for various central nervous system diseases with an inflammatory component. Previously we have identified a small hybrid molecule - MM165 which significantly reduces depressive-like symptoms caused by inflammation induced by lipopolysaccharide administration. In the present study, we investigated whether this compound would mitigate cognitive deficits induced by lipopolysaccharide administration and whether treatment with it would affect the plasma or brain levels of quinolinic acid and kynurenic acid. Neuroinflammation was induced in rats by administering lipopolysaccharide at a dose of 0.5 mg/kg body weight for 10 days. We conducted two tests: novel object recognition and object location, to assess the effect on memory impairment in animals previously treated with lipopolysaccharide. In plasma collected from rats, the concentrations of C-reactive protein and tumor necrosis factor alfa were determined. The concentrations of kynurenic acid and quinolinic acid were determined in plasma and homogenates obtained from the cerebral cortex of rats. Interleukin 6 in the cerebral cortex of rats was determined. Additionally, the body and spleen mass and spontaneous activity were measured in rats. Our study shows that MM165 may mitigate cognitive deficits induced by inflammation after administration of lipopolysaccharide and alter the concentrations of tryptophan metabolites in the brain. Compounds exhibiting a mechanism of action analogous to that of MM165 may serve as foundational structures for the development of a new class of antidepressants.

The protective effects of hesperidin as an antioxidant against quinolinic acid-induced toxicity on oligodendroglia cells: An in vitro study.

INTRODUCTION: Multiple sclerosis (MS) is a complex central nervous system disorder, marked by neurodegenerative and inflammatory processes, where overproduction of reactive oxygen species (ROS) is a key factor in demyelination and neurodegeneration. The current study aims to investigate the effect of hesperidin and Quinolinic acid (QA) on ROS and antioxidant levels, and cell viability of OLN-93 cells. METHODS: OLN-93 cell lines were treated with hesperidin and QA. OLN-93 cells were cultured in Dulbecco's modified Eagle's medium under controlled conditions. Cell viability assays were performed using resazurin to assess the toxicity of hesperidin and QA. Additionally, ROS levels were measured using DCFDA, and malondialdehyde (MDA) levels were determined to evaluate oxidative stress. Superoxide dismutase (SOD) activity and cell viability were assessed by trypan blue staining after exposure to hesperidin and QA. RESULTS: The results of the current study showed that co-administration of 8 mM QA with 50, 100, and 200 µM hesperidin significantly reduced both ROS and MDA levels, demonstrating a substantial attenuation in comparison to the elevated ROS and MDA levels induced by 8 mM QA (p-value < 0.01). Furthermore, 8 mM QA + 50, 100, and 200 µM hesperidin significantly increased SOD levels compared with QA alone (p-value < 0.01). In addition, treatment of OLN cells with 8 mM QA + 50, 100, and 200 µM hesperidin led to higher cell viability compared to QA alone (p value <0.0001). CONCLUSION: The current study demonstrated the antioxidant effect of hesperidin on OLN-93 cells suggesting new insights into the clinical application of hesperidin as an effective treatment for patients with MS. Future in vivo studies, focusing on cellular mechanisms are recommended.

Effect of Quinolinic Acid on Behavior, Morphology, and Expression of Inflammatory/oxidative Status in Rats' Striatum: Is Coenzyme Q10 a Good Protector?

Quinolinic acid (QUIN) is a toxic compound with pro-oxidant, pro-inflammatory, and pro-apoptotic actions found at high levels in the central nervous system (CNS) in several pathological conditions. Due to the toxicity of QUIN, it is important to evaluate strategies to protect against the damage caused by this metabolite in the brain. In this context, coenzyme Q10 (CoQ10) is a provitamin present in the mitochondria with a protective role in cells through several mechanisms of action. Based on these, the present study was aimed at evaluating the possible neuroprotective role of CoQ10 against damage caused by QUIN in the striatum of young Wistar rats. Twenty-one-day-old rats underwent a 10-day pretreatment with CoQ10 or saline (control) intraperitoneal injections and on the 30th day of life received QUIN intrastriatal or saline (control) administration. The animals were submitted to behavior tests or euthanized, and the striatum was dissected to neurochemical studies. Results showed that CoQ10 was able to prevent behavioral changes (the open field, object recognition, and pole test tasks) and neurochemical parameters (alteration in the gene expression of IL-1ß, IL-6, SOD, and GPx, as well as in the immunocontent of cytoplasmic Nrf2 and nuclear p-Nf-κß) caused by QUIN. These findings demonstrate the promising therapeutic effects of CoQ10 against QUIN toxicity.

Bacillus calmette gaurine vaccine ameliorates the neurotoxicity of quinolinic acid in rats via the modulation of antioxidant, inflammatory and apoptotic markers.

A mutation in the Huntingtin gene causes 'Huntington's disease, which presents as a motor and behavioral impairment. Due to the limited drug therapy for this disease, scientists are constantly searching for newer and alternative drugs that may either retard or prevent the progress of the disease. This study aims to explore the neuroprotective potential of Bacillus Calmette Gaurine (BCG) vaccine against quinolinic acid-induced (QA) neurotoxicity in rats. QA (200 nmol/2 µl, i.s) was injected bilaterally into the rat striatum, after which a single dose of BCG (2 × 10^7, cfu) was given to the rats. Animals were assessed for behavioral parameters on the 14th and 21st days. On the 22nd day, animals were sacrificed, brains were harvested, and striatum was separated to evaluate biochemical, inflammatory, and apoptotic mediators. Histopathological studies were performed using Hematoxyline and Eosin staining to assess neuronal morphology. BCG treatment reversed motor abnormalities, reduced oxidative stress and neuroinflammatory markers, apoptotic mediators and striatal lesions induced by QA treatment. In conclusion, treat' 'ing rats with BCG vaccine (2 × 10^7, cfu) mitigated the quinolinic acid-induced Huntington's disease-like symptoms. Hence, BCG vaccine (2 ×10^7, cfu) could be used as an adjuvant in managing HD.

Artemisia Absinthium Extract Attenuates the Quinolinic Acid-Induced Cell Injury in OLN-93 Cells.

OBJECTIVE: Increased quinolinic acid (QA) accumulation has been found in many neurodegenerative diseases. Artemisia absinthium (A. absinthium) has been reported to have neuroprotective and antioxidant activities. This study was designed to evaluate the effect of A. absinthium in QAinduced neurotoxicity in OLN-93 Cells. METHODS: OLN-93 cells were cultured in a DMEM medium containing 10% (v/v) fetal bovine serum, 100 units/ml penicillin, and 100 µg/ml streptomycin. The cells were pretreated with concentrations of A. absinthium extract for two h and then exposed to QA for 24 h. After 24 h cell viability, the level of malondialdehyde (MDA), reactive oxygen species (ROS), and apoptotic cells were quantitated in OLN-93 Cells. RESULTS: Pretreatment with A. absinthium extract prevented the loss of cell viability in OLN-93 cells. ROS generation, lipid peroxidation, and apoptosis in QA-injured OLN-93 cells were reduced following A. absinthium extract pretreatment. CONCLUSION: A. absinthium extract exerts its neuroprotective effect against QA-induced neurotoxicity via oxidative stress and apoptosis modulation.

Exploring the neuroprotective effects of montelukast on brain inflammation and metabolism in a rat model of quinolinic acid-induced striatal neurotoxicity.

BACKGROUND: One intrastriatal administration of quinolinic acid (QA) in rats induces a lesion with features resembling those observed in Huntington's disease. Our aim is to evaluate the effects of the cysteinyl leukotriene receptor antagonist montelukast (MLK), which exhibited neuroprotection in different preclinical models of neurodegeneration, on QA-induced neuroinflammation and regional metabolic functions. METHODS: The right and left striatum of Sprague Dawley and athymic nude rats were injected with QA and vehicle (VEH), respectively. Starting from the day before QA injection, animals were treated with 1 or 10 mg/kg of MLK or VEH for 14 days. At 14 and 30 days post-lesion, animals were monitored with magnetic resonance imaging (MRI) and positron emission tomography (PET) using [18F]-VC701, a translocator protein (TSPO)-specific radiotracer. Striatal neuroinflammatory response was measured post-mortem in rats treated with 1 mg/kg of MLK by immunofluorescence. Rats treated with 10 mg/kg of MLK also underwent a [18F]-FDG PET study at baseline and 4 months after lesion. [18F]-FDG PET data were then used to assess metabolic connectivity between brain regions by applying a covariance analysis method. RESULTS: MLK treatment was not able to reduce the QA-induced increase in striatal TSPO PET signal and MRI lesion volume, where we only detected a trend towards reduction in animals treated with 10 mg/kg of MLK. Post-mortem immunofluorescence analysis revealed that MLK attenuated the increase in striatal markers of astrogliosis and activated microglia in the lesioned hemisphere. We also found a significant increase in a marker of anti-inflammatory activity (MannR) and a trend towards reduction in a marker of pro-inflammatory activity (iNOS) in the lesioned striatum of MLK-compared to VEH-treated rats. [18F]-FDG uptake was significantly reduced in the striatum and ipsilesional cortical regions of VEH-treated rats at 4 months after lesion. MLK administration preserved glucose metabolism in these cortical regions, but not in the striatum. Finally, MLK was able to counteract changes in metabolic connectivity and measures of network topology induced by QA, in both lesioned and non-lesioned hemispheres. CONCLUSIONS: Overall, MLK treatment produced a significant neuroprotective effect by reducing neuroinflammation assessed by immunofluorescence and preserving regional brain metabolism and metabolic connectivity from QA-induced neurotoxicity in cortical and subcortical regions.

Quinolinic Acid Induces Alterations in Neuronal Subcellular Compartments, Blocks Autophagy Flux and Activates Necroptosis and Apoptosis in Rat Striatum.

Quinolinic acid (QUIN) is an agonist of N-methyl-D-aspartate receptor (NMDAr) used to study the underlying mechanism of excitotoxicity in animal models. There is evidence indicating that impairment in autophagy at early times contributes to cellular damage in excitotoxicity; however, the status of autophagy in QUIN model on day 7 remains unexplored. In this study, the ultrastructural analysis of subcellular compartments and the status of autophagy, necroptosis, and apoptosis in the striatum of rats administered with QUIN (120 nmol and 240 nmol) was performed on day 7. QUIN induced circling behavior, neurodegeneration, and cellular damage; also, it promoted swollen mitochondrial crests, spherical-like morphology, and mitochondrial fragmentation; decreased ribosomal density in the rough endoplasmic reticulum; and altered the continuity of myelin sheaths in axons with separation of the compact lamellae. Furthermore, QUIN induced an increase and a decrease in ULK1 and p-70-S6K phosphorylation, respectively, suggesting autophagy activation; however, the increased microtubule-associated protein 1A/1B-light chain 3-II (LC3-II) and sequestosome-1/p62 (SQSTM1/p62), the coexistence of p62 and LC3 in the same structures, and the decrease in Beclin 1 and mature cathepsin D also indicates a blockage in autophagy flux. Additionally, QUIN administration increased tumor necrosis factor alpha (TNFα) and receptor-interacting protein kinase 3 (RIPK3) levels and its phosphorylation (p-RIPK3), as well as decreased B-cell lymphoma 2 (Bcl-2) and increased Bcl-2-associated X protein (Bax) levels and c-Jun N-terminal kinase (JNK) phosphorylation, suggesting an activation of necroptosis and apoptosis, respectively. These results suggest that QUIN activates the autophagy, but on day 7, it is blocked and organelle and cellular damage, neurodegeneration, and behavior alterations could be caused by necroptosis and apoptosis activation.

Quinolinic Acid Impairs Redox Homeostasis, Bioenergetic, and Cell Signaling in Rat Striatum Slices: Prevention by Coenzyme Q10.

Quinolinic acid (QUIN) is an important agonist of NMDA receptors that are found at high levels in cases of brain injury and neuroinflammation. Therefore, it is necessary to investigate neuroprotection strategies capable of neutralizing the effects of the QUIN on the brain. Coenzyme Q10 (CoQ10) is a provitamin that has an important antioxidant and anti-inflammatory action. This work aims to evaluate the possible neuroprotective effect of CoQ10 against the toxicity caused by QUIN. Striatal slices from 30-day-old Wistar rats were preincubated with CoQ10 25-100 µM for 15 min; then, QUIN 100 µM was added to the incubation medium for 30 min. A dose-response curve was used to select the CoQ10 concentration to be used in the study. Results showed that QUIN caused changes in the production of ROS, nitrite levels, activities of antioxidant enzymes, glutathione content, and damage to proteins and lipids. CoQ10 was able to prevent the effects caused by QUIN, totally or partially, except for damage to proteins. QUIN also altered the activities of electron transport chain complexes and ATP levels, and CoQ10 prevented totally and partially these effects, respectively. CoQ10 prevented the increase in acetylcholinesterase activity, but not the decrease in the activity of Na+,K+-ATPase caused by QUIN. We also observed that QUIN caused changes in the total ERK and phospho-Akt content, and these effects were partially prevented by CoQ10. These findings suggest that CoQ10 may be a promising therapeutic alternative for neuroprotection against QUIN neurotoxicity.

Neuroprotective Effects of a Wnt Antagonist in Quinolinic Acid-Induced Excitotoxicity in N18D3 Cells.

Excessive stimulation of the quinolinic acid induces neuronal cell death and is implicated in developing several neurodegenerative diseases. This study investigated whether a Wnt5a antagonist plays a neuroprotective role by regulating the Wnt pathway, activating cellular signaling mechanisms, including MAP kinase and ERK, and acting on the antiapoptotic and the proapoptotic genes in N18D3 neural cells. The cells were pretreated with a Wnt5a antagonist Box5, for one hour and then exposed to quinolinic acid (QUIN), an NMDA receptor agonist for 24 hours. An MTT assay and DAPI staining were used to evaluate cell viability and apoptosis, respectively, demonstrating that Box5 protected the cells from apoptotic death. In addition, a gene expression analysis revealed that Box5 prevented the QUIN-induced expression of the pro-apoptotic genes, BAD and BAX, and increased that of the anti-apoptotic genes, Bcl-xL, BCL2, and BCLW. Further examination of potential cell signaling candidates involved in this neuroprotective effect showed that the immunoreactivity of ERK was significantly increased in the cells treated with Box5. These results suggest that the neuroprotective mechanism of Box5 against QUIN-induced excitotoxic cell death involves the regulation of ERK and modulation of cell survival and death genes through decreasing the Wnt pathway, specifically Wnt5a.

Increased susceptibility to quinolinic acid-induced seizures and long-term changes in brain oscillations in an animal model of glutaric acidemia type I.

Glutaric acidemia type I (GA-I) is an inborn error of metabolism of lysine, hydroxylysine, and tryptophan, caused by glutaryl-CoA-dehydrogenase (GCDH) deficiency, characterized by the buildup of toxic organic acids predominantly in the brain. After acute catabolic states, patients usually develop striatal degeneration, but the mechanisms behind this damage are still unknown. Quinolinic acid (QA), a metabolite of the kynurenine pathway, increases especially during infections/inflammatory processes, and could act synergically with organic acids, contributing to the neurological features of GA-I. The aim of this study was to investigate whether QA increases seizure susceptibility and modifies brain oscillation patterns in an animal model of GA-I, the Gcdh-/- mice taking high-lysine diet (Gcdh-/- -Lys). Therefore, the characteristics of QA-induced seizures and changes in brain oscillatory patterns were evaluated by video-electroencephalography (EEG) analysis recorded in Gcdh-/- -Lys, Gcdh+/+ -Lys, and Gcdh-/- -N (normal diet) animals. We found that the number of seizures per animal was similar for all groups receiving QA, Gcdh-/- -Lys-QA, Gcdh+/+ -Lys-QA, and Gcdh-/- -N-QA. However, severe seizures were observed in the majority of Gcdh-/- -Lys-QA mice (82%), and only in 25% of Gcdh+/+ -Lys-QA and 44% of Gcdh-/- -N-QA mice. All Gcdh-/- -Lys animals developed spontaneous recurrent seizures (SRS), but Gcdh-/- -Lys-QA animals had increased number of SRS, higher mortality rate, and significant predominance of lower frequency oscillations on EEG. Our results suggest that QA plays an important role in the neurological features of GA-I, as Gcdh-/- -Lys mice exhibit increased susceptibility to intrastriatal QA-induced seizures and long-term changes in brain oscillations.

Mast cells and histamine are involved in the neuronal damage observed in a quinolinic acid-induced model of Huntington's disease.

Huntington´s disease (HD) is a pathological condition that can be studied in mice by the administration of quinolinic acid (QUIN), an agonist of the N-methyl-d-aspartate receptor (NMDAR) that induces NMDAR-mediated cytotoxicity and neuroinflammation. Mast cells (MCs) participate in numerous inflammatory processes through the release of important amounts of histamine (HA). In this study, we aimed to characterize the participation of MCs and HA in the establishment of neural and oxidative damage in the QUIN-induced model of HD. C57BL6/J mice (WT), MC-deficient c-KitW-sh/W-sh (Wsh) mice and Wsh mice reconstituted by intracerebroventricular (i.c.v.) injection of 5 × 105 bone marrow-derived mast cells (BMMCs), or i.c.v. administered with HA (5 µg) were used. All groups of animals were intrastriatally injected with 1 µL QUIN (30 nmol/µL) and 3 days later, apomorphine-induced circling behavior, striatal GABA levels and the number of Fluoro-Jade positive cells, as indicators of neuronal damage, were determined. Also, lipid peroxidation (LP) and reactive oxygen species production (ROS), as markers of oxidative damage, were analyzed. Wsh mice showed less QUIN-induced neuronal and oxidative damage than WT and Wsh-MC reconstituted animals. Histamine administration restored the QUIN-induced neuronal and oxidative damage in the non-reconstituted Wsh mice to levels equivalent or superior to those observed in WT mice. Our results demonstrate that MCs and HA participate in the neuronal and oxidative damages observed in mice subjected to the QUIN -induced model of Huntington's disease.

5,6,7 trihydroxy flavone armoured neurodegeneration caused by Quinolinic acid induced huntington's like disease in rat striatum - reinstating the level of brain neurotrophins with special reference to cognitive-socio behaviour, biochemical and histopathological aspects.

Huntington Disease (HD), a predominant Neurodegenerative Disorder which might be induced by endogenous neurotoxin called Quinolinic Acid (QA), an N-methyl-D aspartate receptor (NMDAR) agonist, the bilaterally intrastriatal administration (200 nm/2 µL of saline) offers rise to the toxic events like neuronal death, neuroinflammation by inflicting excitotoxicity and oxidative stress in the striatum of male Wistar rats by exhibiting the behavioural changes which was accessed by rotarod, open field analysis. In this study, the neuropharmacological effect of Baicalein (BC) against QA induced HD was evaluated. Baicalein (BC), scientifically 5,6,7 trihydroxy flavone present naturally in the edible plants like Scutellaria baicalensis and Oroxylum indicum possess a better neuroprotective effect in the dosage of 10 mg/kg and 30 mg/kg intraperitoneally in the striatum of HD induced rats. This study proved that BC is efficient to revive the level of enzymatic & non-enzymatic antioxidants and mitochondrial complexes by decreasing the number of inflammatory mediators such as MDA, protein carbonyls and Nitric Oxide at the significance of P < 0.01 and restores the amount of BDNF and GDNF thereby preventing the neurophysiological changes which were analysed by haematoxylin & eosin staining. Thus finally, the protective effect of Baicalein displays the up-gradation of psychological and behavioural changes induced by QA.

Kynurenic acid in neurodegenerative disorders-unique neuroprotection or double-edged sword?

AIMS: The family of kynurenine pathway (KP) metabolites includes compounds produced along two arms of the path and acting in clearly opposite ways. The equilibrium between neurotoxic kynurenines, such as 3-hydroxykynurenine (3-HK) or quinolinic acid (QUIN), and neuroprotective kynurenic acid (KYNA) profoundly impacts the function and survival of neurons. This comprehensive review summarizes accumulated evidence on the role of KYNA in Alzheimer's, Parkinson's and Huntington's diseases, and discusses future directions of potential pharmacological manipulations aimed to modulate brain KYNA. DISCUSSION: The synthesis of specific KP metabolites is tightly regulated and may considerably vary under physiological and pathological conditions. Experimental data consistently imply that shift of the KP to neurotoxic branch producing 3-HK and QUIN formation, with a relative or absolute deficiency of KYNA, is an important factor contributing to neurodegeneration. Targeting specific brain regions to maintain adequate KYNA levels seems vital; however, it requires the development of precise pharmacological tools, allowing to avoid the potential cognitive adverse effects. CONCLUSIONS: Boosting KYNA levels, through interference with the KP enzymes or through application of prodrugs/analogs with high bioavailability and potency, is a promising clinical approach. The use of KYNA, alone or in combination with other compounds precisely influencing specific populations of neurons, is awaiting to become a significant therapy for neurodegenerative disorders.

Non-invasive, neurotoxic surgery reduces seizures in a rat model of temporal lobe epilepsy.

Surgery can be highly effective for treating certain cases of drug resistant epilepsy. The current study tested a novel, non-invasive, surgical strategy for treating seizures in a rat model of temporal lobe epilepsy. The surgical approach uses magnetic resonance-guided, low-intensity focused ultrasound (MRgFUS) in combination with intravenous microbubbles to open the blood-brain barrier (BBB) in a transient and focal manner. During the period of BBB opening, a systemically administered neurotoxin (Quinolinic Acid: QA) that is normally impermeable to the BBB gains access to a targeted area in the brain, destroying neurons where the BBB has been opened. This strategy is termed Precise Intracerebral Non-invasive Guided Surgery (PING). Spontaneous recurrent seizures induced by pilocarpine were monitored behaviorally prior to and after PING or under control conditions. Seizure frequency in untreated animals or animals treated with MRgFUS without QA exhibited expected seizure rate fluctuations frequencies between the monitoring periods. In contrast, animals treated with PING targeting the intermediate-temporal aspect of the hippocampus exhibited substantial reductions in seizure frequency, with convulsive seizures being eliminated entirely in two animals. These findings suggest that PING could provide a useful alternative to invasive surgical interventions for treating drug resistant epilepsy, and perhaps for treating other neurological disorders in which aberrant neural circuitries play a role.

Papaverine, a Phosphodiesterase 10A Inhibitor, Ameliorates Quinolinic Acid-Induced Synaptotoxicity in Human Cortical Neurons.

Phosphodiesterase-10A (PDE10A) hydrolyse the secondary messengers cGMP and cAMP, two molecules playing important roles in neurodevelopment and brain functions. PDE10A is associated to progression of neurodegenerative diseases like Alzheimer's, Parkinson's, Huntington's diseases, and a critical role in cognitive functions. The present study was undertaken to determine the possible neuroprotective effects and the associated mechanism of papaverine (PAP), a PDE10A isoenzyme inhibitor, against quinolinic acid (QUIN)-induced excitotoxicity using human primary cortical neurons. Cytotoxicity potential of PAP was analysed using MTS assay. Reactive oxygen species (ROS) and mitochondrial membrane potential were measured by DCF-DA and JC10 staining, respectively. Caspase 3/7 and cAMP levels were measured using ELISA kits. Effect of PAP on the CREB, BNDF and synaptic proteins such as SAP-97, synaptophysin, synapsin-I, and PSD-95 expression was analysed by Western blot. Pre-treatment with PAP increased intracellular cAMP and nicotinamide adenine dinucleotide (NAD+) levels, restored mitochondrial membrane potential (ΔΨm), and decreased ROS and caspase 3/7 content in QUIN exposed neurons. PAP up-regulated CREB and BDNF, and synaptic protein expression. In summary, these data indicate that PDE10A is involved in QUIN-mediated synaptotoxicity and its inhibition elicit neuroprotection by reducing the oxidative stress and protecting synaptic proteins via up-regulation of cAMP signalling cascade.

Kynurenic Acid Protects Against Reactive Glial-associated Reductions in the Complexity of Primary Cortical Neurons.

Brain glia produce neuroactive metabolites via tryptophan-kynurenine catabolism. A role for kynurenine pathway (KP) metabolites is proposed in reactive glial associated neurodegeneration. The aim of this investigation was to assess the role of KP induction and KP metabolites in driving reactive glial associated neuronal atrophy. Rat primary mixed glia, and enriched microglial and astroglial cultures were stimulated with IFNγ (10 ng/ml) for 24 hours. KP induction in mixed glial cells was confirmed by raised expression of the rate limiting KP enzyme indoleamine 2,3 dioxygenase (IDO) and raised concentrations of KP metabolites kynurenic acid (KYNA) and quinolinic acid (QUIN) in the conditioned media. Conditioned media was transferred onto immature (3 days) and mature (21 days) primary cortical neurons in vitro for 24 hours. IFNγ-stimulated mixed glial conditioned media reduced neurite outgrowth and complexity of both immature and mature neurons and co-localised expression of synaptic markers determined by immunocytochemistry. Pre-treatment of mixed glial cells with the IDO inhibitor, 1-methyltryptophan (1-MT) (L) prevented these effects of IFNγ-stimulated mixed glial conditioned media. KYNA increased complexity and synapse formation in mature cortical neurons and protected against reduced neuronal complexity and co-localised expression of synaptic markers induced by conditioned media from IFNγ-stimulated mixed glia and by treatment of neuronal cells with QUIN (1 µM). Overall, this study supports a role for the KP in driving neuronal atrophy associated with reactive glia and indicates that inhibition of the KP in glia, or raising the concentration of the astrocytic metabolite KYNA, protects against reactive microglial and QUIN-associated neuronal atrophy.

Neuroprotective effects of roflumilast against quinolinic acid-induced rat model of Huntington's disease through inhibition of NF-κB mediated neuroinflammatory markers and activation of cAMP/CREB/BDNF signaling pathway.

Huntington's disease (HD) is a progressive neurodegenerative and hyperkinetic movement disorder. Decreased activity of cAMP-responsive element-binding protein (CREB) is thought to contribute to the death of striatal medium spiny neurons in HD. The present study has been designed to explore the possible role of roflumilast against qunilonic acid (QA) induced neurotoxicity in rats intending to investigate whether it inhibits the neuroinflammatory response through activation of the cAMP/CREB/BDNF signaling pathway. QA was microinjected (200 nmol/2 µl, bilaterally) through the intrastriatal route in the stereotaxic apparatus. Roflumilast (0.5, 1, and 2 mg/kg, orally) once-daily treatment for 21 days significantly improved locomotor activity in actophotometer, motor coordination in rotarod, and impaired gait performance in narrow beam walk test. Moreover, roflumilast treatment significantly attenuated oxidative and nitrosative stress (p < 0.05) through attenuating lipid peroxidation nitrite concentration and enhancing reduced glutathione, superoxide dismutase, and catalase levels. Furthermore, roflumilast also significantly decreased elevated pro-inflammatory cytokines like TNF-α (p < 0.01), IL-6 (p < 0.01), IFN-γ (p < 0.05), NF-κB (p < 0.05) and significantly increased BDNF(p < 0.05) in the striatum and cortex of rat brain. The results further demonstrated that roflumilast effectively increased the gene expression of cAMP(p < 0.05), CREB(p < 0.05) and decreased the gene expression of PDE4 (p < 0.05) in qRT-PCR. These results conclusively depicted that roflumilast could be a potential candidate as an effective therapeutic agent in the management of HD through the cAMP/CREB/BDNF signaling pathway.

URB597 Prevents the Short-Term Excitotoxic Cell Damage in Rat Cortical Slices: Role of Cannabinoid 1 Receptors.

Endocannabinoid-based therapies constitute an emerging tool for the potential treatment of neurodegenerative disorders, requiring characterization at the experimental level. The effects of URB597, an inhibitor of the fatty acid amide hydrolase (FAAH), were tested against the quinolinic acid (QUIN)-induced early toxic effects in rat cortical slices, and compared with those effects exerted by the endocannabinoid anandamide (AEA). URB597 prevented the QUIN-induced loss of mitochondrial function/cell viability and lipid peroxidation, while reduced necrosis, and to a lesser extent, apoptosis. The protective effects of URB597 were mediated by activation of cannabinoid receptor 1 (CB1r), as evidenced by their inhibition by the selective CB1r antagonist AM281. Similar effects were observed when testing AEA against QUIN toxicity. Our findings demonstrate the neuroprotective properties of URB597 during the early stages of excitotoxic damage to cortical tissue, suggesting that these properties are mediated by FAAH inhibition, and might be linked to the protective effects of AEA, or the combination of endocannabinoids.

Cerebral dopamine neurotrophic factor (CDNF) protects against quinolinic acid-induced toxicity in in vitro and in vivo models of Huntington's disease.

Huntington's disease (HD) is a neurodegenerative disorder with a progressive loss of medium spiny neurons in the striatum and aggregation of mutant huntingtin in the striatal and cortical neurons. Currently, there are no rational therapies for the treatment of the disease. Cerebral dopamine neurotrophic factor (CDNF) is an endoplasmic reticulum (ER) located protein with neurotrophic factor (NTF) properties, protecting and restoring the function of dopaminergic neurons in animal models of PD more effectively than other NTFs. CDNF is currently in phase I-II clinical trials on PD patients. Here we have studied whether CDNF has beneficial effects on striatal neurons in in vitro and in vivo models of HD. CDNF was able to protect striatal neurons from quinolinic acid (QA)-induced cell death in vitro via increasing the IRE1α/XBP1 signalling pathway in the ER. A single intrastriatal CDNF injection protected against the deleterious effects of QA in a rat model of HD. CDNF improved motor coordination and decreased ataxia in QA-toxin treated rats, and stimulated the neurogenesis by increasing doublecortin (DCX)-positive and NeuN-positive cells in the striatum. These results show that CDNF positively affects striatal neuron viability reduced by QA and signifies CDNF as a promising drug candidate for the treatment of HD.

Longitudinal Magnetic Resonance Spectroscopy and Diffusion Tensor Imaging in Sheep (Ovis aries) With Quinolinic Acid Lesions of the Striatum: Time-Dependent Recovery of N-Acetylaspartate and Fractional Anisotropy.

We created an excitotoxic striatal lesion model of Huntington disease (HD) in sheep, using the N-methyl-d-aspartate receptor agonist, quinolinic acid (QA). Sixteen sheep received a bolus infusion of QA (75 µL, 180 mM) or saline, first into the left and then (4 weeks later) into the right striatum. Magnetic resonance spectroscopy (MRS) and diffusion tensor imaging (DTI) of the striata were performed. Metabolite concentrations and fractional anisotropy (FA) were measured at baseline, acutely (1 week after each surgery) and chronically (5 weeks or greater after the surgeries). There was a significant decrease in the neuronal marker N-acetylaspartate (NAA) and in FA in acutely lesioned striata of the QA-lesioned sheep, followed by a recovery of NAA and FA in the chronically lesioned striata. NAA level changes indicate acute death and/or impairment of neurons immediately after surgery, with recovery of reversibly impaired neurons over time. The change in FA values of the QA-lesioned striata is consistent with acute structural disruption, followed by re-organization and glial cell infiltration with time. Our study demonstrates that MRS and DTI changes in QA-sheep are consistent with HD-like pathology shown in other model species and that the MR investigations can be performed in sheep using a clinically relevant human 3T MRI scanner.